Transplantation is the best therapy for end stage failure of many organs including kidney, heart and lungs. While short term outcomes of transplants are excellent, graft half-lives are suboptimal in large part because of chronic immune mediated injury. Our long term goal is to identify novel mechanisms of transplant injury so as to be able to apply them to develop therapuetic strategies to prolong transplant survival. Increasing evidence indicates that regulatory T cells (Tregs) could be harnessed to inhibit pathogenic anti-transplant immunity (in the absence of immune suppression) but mechanisms to accomplish this goal are hampered by inadequate understanding. Extending our previous work, our combined preliminary data demonstrate that complement components C3a and C5a (initially thought to be important only for innate immunity) signal via their receptors on both mouse and human Treg inhibiting induction, activation, and function whereas blockade markedly augments Treg induction, activation, function. To decipher the mechanistic connections between complement and Treg in transplantation we propose a series of studies using mouse and human cells (in vitro) and multiple in vivo transplant models in mice. We will use knockout and transgenic mice along with pharmacological inhibitors, blocking antibodies and knockdown technologies to a) determine the mechanisms linking C3aR/C5aR signaling with induction and function of murine and human Treg and b) test the in vivo effects of C3aR/C5aR signaling on iTreg and nTreg in transplantation. The expected outcomes are that we will delineate the effects of immune cell C3aR/C5aR signaling on the function of mouse Tregs in transplantation, define the molecular signaling pathways and mechanisms that explain the observed effects, and test how the murine findings translate to human Tregs. Together, the results are likely to identify novel and unanticipated immune mechanisms (innovation), and have a positive impact because they will provide new, mechanistic insight into Treg function and stability, and identify new therapeutic targets that have the potential to prolong transplant survival in humans.